Improving a surrogate eggshell incubation system using white leghorn chicken eggs obtained from a poultry flock of the same age

Abstract A germinal disc located on egg yolk surface drives genetic modification. Based on eggshells, in ovo incubation such as windowed and surrogate methods have been developed, but these exhibit limited abilities to generate transgenic chickens. Here, we investigated the frequency of observing the germinal disc according to the preincubation positioning direction and time and found that it depended on those conditions, but only a few chicks (2.8-5.6%) hatched using windowed method. Then, we attempted to improve surrogate method via one- or two-step procedures, and all eggs, including surrogates which is 10 g heavier than donor egg, were obtained from a poultry flock of the same age. With the one-step surrogate method (the donor egg is transferred only once into the surrogate egg), the survival rate at day 4 was 30.8%, and the hatching rate was 11.8%. We then established an improved two-step surrogate method: System II is similar with methods previously established, and an unusual second surrogate egg is used in System III. With this method, the survival rate at day 4 was improved by 90.7%, and a hatching rate of 70.0% was obtained. Therefore, the improved surrogate methods established in this study can be effective for in ovo incubation.